Hemophilia A is a severe hemorrhagic disorder due to deficiency of coagulation factor VIII. Approximately 15 to 20 percent of severely affected patients develop antibodies to factor VIII with replacement therapy. These antibodies, termed inhibitors, neutralize factor VIII therapy causing significant morbidity and mortality. Recent studies in animal models suggest that inhibitors are also likely to hinder the success of gene therapy approaches. Several lines of evidence, including analyses of hemophilic brother pairs and animal models, indicate that the propensity to develop inhibitors is strongly influenced by genetic factors other than the specific factor VIII mutation or histocompatibility (MHC) locus type. This proposal aims to employ newly developed techniques for genetic linkage analysis and genome scanning to identify the inhibitor susceptibility genes. At least 200 well characterized severe hemophilia A brother pairs, both concordant and discordant for inhibitor formation, and other informative family members will be identified through a collaborative network of investigators in over 70 hemophilia treatment centers. Samples will be collected, and a repository of genomic DNA accompanied by a clinical and epidemiologic data base characterizing the hemophilic inhibitor phenotype including screening for the intron 22 inversion mutation, will be created. A panel of candidate genes that are likely to play a positive or negative role in inhibitor development will then be examined through fine mapping of polymorphic markers in the regions of interest, including the MHC, immunoglobulin genes, the T-cell receptor genes, and selected cytokine and cytokine receptor genes. If candidate genes fail to account for the genetic susceptibility to inhibitor development, a total 10 cM genome scan utilizing approximately 350 highly polymorphic microsatellite repeat markers will be performed. Linkage analysis will include both non-parametric testing of brother pairs for multiple susceptibility genes and more powerful parametric methods for one and two gene models in extended pedigrees. Identification of new genomic regions linked to inhibitor development will then be explored with fine mapping of the chromosomal areas and gene cloning techniques. The DNA bank and database will also be a uniquely valuable resource for future studies of patients with hemophilia.